1.山西医科大学,太原 030001;2.山西医科大学第一医院 消化内科,太原 030001
R-33
1. Shanxi Medical University, Taiyuan 030001, China. 2. Gastroenterology Department, the First Hospital of Shanxi Medical University, Taiyuan 030001
目的 研究肝星状细胞活化及增殖过程中还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶4(NOX4)和信号转导与转录激活因子3(STAT3)表达的变化,并探讨NOX4 抑制剂GKT137831 是否可通过抑制NOX4,减少活性氧(ROS)的产生,抑制STAT3 信号通路,抑制肝星状细胞活化与增殖。 方法 将大鼠肝星状细胞-T6(HSC-T6)分为4 组:对照组、TGF-β1 组、TGF-β1+GKT137831 组、GKT137831 组。DCFH-DA 荧光探针法检测HSC-T6 细胞内ROS 水平,CCK-8 法检测细胞增殖,实时荧光定量PCR 法检测NOX4、STAT3 mRNA 的表达;细胞免疫荧光和蛋白印迹法检测NOX4、STAT3、p-STAT3 及α-SMA 蛋白的表达。 结果 与对照组相比,TGF-β1 组HSC-T6 细胞中NOX4mRNA 和蛋白表达上调,细胞内ROS 水平升高,p-STAT3 蛋白表达增加,细胞活化增殖能力升高(P<0. 05);给予GKT137831 处理后,HSC-T6 细胞中NOX4 mRNA 和蛋白表达下调,细胞内ROS 水平下降,STAT3 蛋白磷酸化减少,细胞活化增殖能力下降(P<0. 05)。 结论 NOX4 可能通过产生ROS,促进STAT3 磷酸化,导致HSC-T6 细胞活化与增殖;GKT137831 通过抑制NOX4 来减少ROS 的产生,抑制STAT3 磷酸化,从而抑制HSC-T6 细胞活化与增殖。
Objective To investigate changes to the expression of reduced nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) and signal transducer and activator of transcription 3 (STAT3) during the activation and proliferation of hepatic stellate cells and to explore whether the NOX4 inhibitor GKT137831 can reduce the production of reactive oxygen species (ROS), as well as inhibit the STAT3 signaling pathway and the activation and proliferation of hepatic stellate cells. Methods Rat hepatic stellate cells-T6 (HSCs-T6) were divided into four groups: control group, TGF-β1 group, TGF-β1+GKT137831 group, and GKT137831 group. The ROS levels in HSC-T6 cells were detected by DCFH-DA fluorescent probe method . The CCK-8 method was used to detect cell proliferation, and real-time fluorescent quantitative PCR was used to detect the expression of NOX4 and STAT3 mRNA. The expression of NOX4, STAT3, p-STAT3, and α-SMA proteins was detected by immunofluorescence and Western blot. Results Compared with the control group, HSC-T6 cells in the TGF-β1 group’s expression of NOX4 mRNA and protein were up-regulated, and intracellular ROS levels, expression of p-STAT3 protein, and the cells’ activation and proliferation ability increased (P< 0. 05). After addition of GKT137831, NOX4 mRNA and protein expressions were down-regulated in HSC-T6 cells, and intracellular ROS levels, STAT3 protein phosphorylation, and the cells’ activation and proliferation ability were decreased (P<0. 05). Conclusions NOX4 may promote the phosphorylation of STAT3 by producing ROS, leading to the activation and proliferation of HSC-T6 cells. By inhibiting NOX4, GKT137831 reduces ROS production and inhibits STAT3 phosphorylation, thereby inhibiting HSC-T6 cell activation and proliferation.
孟妍芳. NOX4/ ROS/ STAT3 通路对肝星状细胞活化增殖的影响[J].中国比较医学杂志,2023,33(4):90~95.
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